A SQUAMOSA MADS-box gene involved in the regulation of anthocyanin accumulation in bilberry fruits

Anthocyanins are important health promoting phytochemicals that are abundant in many fleshy fruits. Bilberry (Vaccinium myrtillus L.) is one of the best sources of these compounds. Here we report on the expression pattern and functional analysis of a SQUAMOSA (SQUA) class MADS-box transcription factor, VmTDR4, associated with anthocyanin biosynthesis in bilberry. Levels of VmTDR4 expression were spatially and temporally linked with colour development and anthocyanin-related gene expression. Virus induced gene silencing (VIGS) was used to suppress VmTDR4 expression in bilberry resulting in substantial reduction in anthocyanin levels in fully ripe fruits. Chalcone synthase was used a positive control in the VIGS experiments. Additionally, in sectors of fruit tissue in which the expression of the VmTDR4 gene was silenced, the expression of R2R3 MYB family transcription factors related to the biosynthesis of flavonoids were also altered. We conclude that VmTDR4 plays an important role in the accumulation of anthocyanins during normal ripening in bilberry; probably through direct or indirect control of transcription factors belonging to the R2R3 MYB family

The meristem-associated endosymbiont Methylorubrum extorquens DSM13060 reprograms development and stress responses of pine seedlings

Microbes living in plant tissues-endophytes-are mainly studied in crop plants where they typically colonize the root apoplast. Trees-a large carbon source with a high capacity for photosynthesis-provide a variety of niches for endophytic colonization. We have earlier identified a new type of plant-endophyte interaction in buds of adult Scots pine, where Methylorubrum species live inside the meristematic cells. The endosymbiont Methylorubrum extorquens DSM13060 significantly increases needle and root growth of pine seedlings without producing plant hormones, but by aggregating around host nuclei. Here, we studied gene expression and metabolites of the pine host induced by M. extorquens DSM13060 infection. Malic acid was produced by pine to potentially boost M. extorquens colonization and interaction. Based on gene expression, the endosymbiont activated the auxin- and ethylene (ET)-associated hormonal pathways through induction of CUL1 and HYL1, and suppressed salicylic and abscisic acid signaling of pine. Infection by the endosymbiont had an effect on pine meristem and leaf development through activation of GLP1-7 and ALE2, and suppressed flowering, root hair and lateral root formation by downregulation of AGL8, plantacyanin, GASA7, COW1 and RALFL34. Despite of systemic infection of pine seedlings by the endosymbiont, the pine genes CUL1, ETR2, ERF3, HYL, GLP1-7 and CYP71 were highly expressed in the shoot apical meristem, rarely in needles and not in stem or root tissues. Low expression of MERI5, CLH2, EULS3 and high quantities of ononitol suggest that endosymbiont promotes viability and protects pine seedlings against abiotic stress. Our results indicate that the endosymbiont positively affects host development and stress tolerance through mechanisms previously unknown for endophytic bacteria, manipulation of plant hormone signaling pathways, downregulation of senescence and cell death-associated genes and induction of ononitol biosynthesis.Peer reviewe

Alcohol intoxication and lack of helmet use are common in electric scooter-related traumatic brain injuries: a consecutive patient series from a tertiary university hospital

Purpose: Clinicians have increasingly encountered traumatic brain injuries (TBI) related to electric scooter (ES) accidents. In this study, we aim to identify the modifiable risk factors for ES-related TBIs.Methods: A retrospective cohort of consecutive patients treated for ES-related traumatic brain injuries in a tertiary university hospital between May 2019 and September 2021 was identified and employed for the study. The characteristics of the accidents along with the clinical and imaging findings of the injuries were collected from the patient charts.Results: During the study period, 104 TBIs related to ES accidents were identified. There was a high occurrence of accidents late at night and on Saturdays. In four cases, the patient’s helmet use was mentioned (3.8%). Seventy-four patients (71%) were intoxicated. At the scene of the accident, seventy-seven (74%) of the patients had a Glasgow Coma Scale score of 13–15, three patients (3%) had a score of 9–12, and two patients (2%) had a score of 3–8. The majority (83%) of TBIs were diagnosed as concussions. Eighteen patients had evidence of intracranial injuries in the imagining. Two patients required neurosurgical procedures. The estimated population standardized incidence increased from 7.0/100,000 (95% CI 3.5–11/100,000) in 2019 to 27/100,000 (95% CI 20–34/100,000) in 2021.Conclusions: Alcohol intoxication and the lack of a helmet were common in TBIs caused by ES accidents. Most of the accidents occurred late at night. Targeting these modifiable factors could decrease the incidence of ES-related TBIs.</p

Alcohol intoxication and lack of helmet use are common in electric scooter-related traumatic brain injuries: a consecutive patient series from a tertiary university hospital

Purpose: Clinicians have increasingly encountered traumatic brain injuries (TBI) related to electric scooter (ES) accidents. In this study, we aim to identify the modifiable risk factors for ES-related TBIs.Methods: A retrospective cohort of consecutive patients treated for ES-related traumatic brain injuries in a tertiary university hospital between May 2019 and September 2021 was identified and employed for the study. The characteristics of the accidents along with the clinical and imaging findings of the injuries were collected from the patient charts.Results: During the study period, 104 TBIs related to ES accidents were identified. There was a high occurrence of accidents late at night and on Saturdays. In four cases, the patient’s helmet use was mentioned (3.8%). Seventy-four patients (71%) were intoxicated. At the scene of the accident, seventy-seven (74%) of the patients had a Glasgow Coma Scale score of 13–15, three patients (3%) had a score of 9–12, and two patients (2%) had a score of 3–8. The majority (83%) of TBIs were diagnosed as concussions. Eighteen patients had evidence of intracranial injuries in the imagining. Two patients required neurosurgical procedures. The estimated population standardized incidence increased from 7.0/100,000 (95% CI 3.5–11/100,000) in 2019 to 27/100,000 (95% CI 20–34/100,000) in 2021.Conclusions: Alcohol intoxication and the lack of a helmet were common in TBIs caused by ES accidents. Most of the accidents occurred late at night. Targeting these modifiable factors could decrease the incidence of ES-related TBIs.</p

Fungal Dysbiosis and Intestinal Inflammation in Children With Beta-Cell Autoimmunity

Although gut bacterial dysbiosis is recognized as a regulator of beta-cell autoimmunity, no data is available on fungal dysbiosis in the children at the risk of type 1 diabetes (T1D). We hypothesized that the co-occurrence of fungal and bacterial dysbiosis contributes to the intestinal inflammation and autoimmune destruction of insulin-producing beta-cells in T1D. Fecal and blood samples were collected from 26 children tested positive for at least one diabetes-associated autoantibody (IAA, GADA, IA-2A or ICA) and matched autoantibody-negative children with HLA-conferred susceptibility to T1D (matched for HLA-DQB1 haplotype, age, gender and early childhood nutrition). Bacterial 16S and fungal ITS2 sequencing, and analyses of the markers of intestinal inflammation, namely fecal human beta-defensin-2 (HBD2), calprotectin and secretory total IgA, were performed. Anti-Saccharomyces cerevisiae antibodies (ASCA) and circulating cytokines, IFNG, IL-17 and IL-22, were studied. After these analyses, the children were followed for development of clinical T1D (median 8 years and 8 months). Nine autoantibody positive children were diagnosed with T1D, whereas none of the autoantibody negative children developed T1D during the follow-up. Fungal dysbiosis, characterized by high abundance of fecal Saccharomyces and Candida, was found in the progressors, i.e., children with beta-cell autoimmunity who during the follow-up progressed to clinical T1D. These children showed also bacterial dysbiosis, i.e., increased Bacteroidales and Clostridiales ratio, which was, however, found also in the non-progressors, and is thus a common nominator in the children with beta-cell autoimmunity. Furthermore, the progressors showed markers of intestinal inflammation detected as increased levels of fecal HBD2 and ASCA IgG to fungal antigens. We conclude that the fungal and bacterial dysbiosis, and intestinal inflammation are associated with the development of T1D in children with beta-cell autoimmunity

The interaction between the intracellular endophytic bacterium, Methylobacterium extorquens DSM13060, and Scots pine (Pinus sylvestris L.)

Abstract To date, plant endophytic bacteria have mainly been studied in roots of crop plants. However, shoot-associated endophytes are less diverse than root-associated ones. Hence, endophytic bacteria of plant shoots evolved different traits, than root colonizers, especially with types of host tissues infected and patterns of growth and development. This study found Methylobacterium extorquens colonized pine seedlings similarly to stem-colonizing rhizobia of other plants. M. extorquens DSM13060 was isolated from meristematic cells in shoot tip cultures of Scots pine (Pinus sylvestris L.). M. extorquens infected the plant stem through epidermis or stomatal apertures, forming infection pockets in the root and stem epidermis, or cortex. Post-infection, thread-like infection structures passed through the endoderm, invading vascular tissues. This led to systemic colonization of above and below ground-parts, observed in in vitro grown Scots pine. A novel mechanism enabling development of endophyte-host symbiosis is discovered within the M. extorquens – Scots pine model. This mechanism involves ability of M. extorquens to produce polyhydroxybutyrates (PHB) to protect itself from host-induced oxidative stress during infection. Upon initial colonization on the host surface, M. extorquens DSM13060 consumes methanol as a carbon source, using it to biosynthesize PHB. PHB are then degraded, upon host infection, by PHB depolymerases (PhaZ) to yield methyl-esterified 3-hydroxybutyrate oligomers. These oligomers have substantial antioxidant activity towards host-induced oxidative stress, enabling the bacterium to bypass host defenses and colonize further tissues. The bacteria can also store PHBs for future protection. The capacity for PHB production and, thus, protection from oxidative stress, is discovered in a wide taxonomic range of bacteria. This study also shows meristematic endophytes are important in growth and development of their hosts. Unlike many bacterial root endophytes, M. extorquens DSM13060 does not induce plant growth through hormones. However, this bacterium can colonize the interior of living host cells, where it aggregates around the nucleus of the host plant. M. extorquens DSM13060 genome encodes nucleomodulins, eukaryotic-like transcription factors, which may intervene in host transcription and metabolism.Tiivistelmä Kasvin sisällä elävien endofyyttisten bakteerien tutkimus on perinteisesti keskittynyt viljelykasveihin ja niiden juuristoon. Kasvien maanpäällisissä versoissa elävät endofyytit eroavat merkittävästi juuriston bakteereista lajirikkauden suhteen. Versoissa eläville bakteereille on todennäköisesti kehittynyt erilaisia sopeumia kuin juuriston endofyyttilajeille. Endofyyttinen Methylobacterium extorquens DSM13060 elää männyn silmujen kasvusolukossa lisäten isäntäkasvin kasvua. Tässä tutkimuksessa M. extorquens –bakteerin todettiin siirtyvän männyn taimiin samoja mekanismeja käyttäen kuin Rhizobium –suvun typensitojabakteerit. Metylobakteeri tunkeutui isäntäkasviin aktiivisesti soluseinien läpi tai varren ilmarakojen kautta muodostaen mikropesäkkeitä juuren ja varren pinnoille, sekä infektiotaskuja kuorisolukkoon. Bakteeri eteni infektiolankojen avulla endodermin ohi johtosolukoihin, mikä mahdollisti bakteerin siirtymisen muualle taimeen. M. extorquens käytti kasvin pinnalla runsaana olevaa metanolia hiilenlähteenään, varastoiden sen solujen sisäiseksi polyhydroksibutyraatti (PHB) polymeeriksi. Infektion myöhemmissä vaiheissa bakteeri hajotti varastoidun polymeerin PHB-depolymeraasientsyymien (PhaZ) avulla lyhyiksi rasvahappoketjuiksi. Nämä metyloidut 3-hydroksibutyraatin oligomeerit suojasivat bakteeria isäntäkasvin puolustuksen tuottamilta happiradikaaleilta mahdollistaen infektion etenemisen. Tutkimuksessa saatujen tulosten perusteella endofyytin solunsisäinen energiavarasto, PHB, toimii pelkistävänä varastona ympäristön hapettavaa stressiä vastaan. Löytö osoitti uudenlaisen antioksidatiivisen puolustumekanismin, joka on levinnyt laajalle bakteerikunnassa ja liittyy yleisesti bakteerien kykyyn sietää vaikeita olosuhteita. Toisin kuin useat juurissa elävät bakteeriendofyytit, M. extorquens ei lisää isäntäkasvin kasvua tuottamalla kasvihormoneja. Bakteeri kykenee elämään männyn elävien solujen sisällä tumien läheisyydessä. M. extorquens DSM13060 genomi sisältääkin useita geenejä, jotka koodaavat nukleomoduliineja, eukaryoottisolujen säätylytekijöiden kaltaisia entsyymejä, joiden avulla bakteeri todennäköisesti vaikuttaa isäntäkasvin aineenvaihduntaan. Vastaavaa vaikutusmekanismia ei ole aikaisemmin kuvattu endofyyteillä. Tutkimus korostaa aiemmin tuntemattomien meristemaattisten bakteeriendofyyttien merkitystä isäntäkasvin kasvussa ja erilaistumisessa

Corpse management of the invasive Argentine ant inhibits growth of pathogenic fungi

Abstract A dead conspecific poses a potential pathogen risk for social animals. We have discovered that Argentine ants (Linepithema humile) prevent spread of pathogenic fungi from corpses by depositing the dead to combined toilet and refuse areas and applying pygidial gland secretion on them. The presence of a corpse in a nest increases this secretion behaviour. We identified three fungi growing on Argentine ant corpses. Growth of the Argentine ant pathogen Aspergillus nomius and the plant pathogen Fusarium solani on corpses was inhibited as long as the ants were constantly attending them as the ant anal secretion only delayed germination of their spores. In contrast, the effect of the ant anal secretion on the human pathogen Aspergillus fumigatus was much stronger: it prevented spore germination and, accordingly, the fungus no longer grew on the treated corpses. The Argentine ants are one of the world’s worst invasive alien species as they cause ecological and economical damage in their new habitats. Our discovery points at a novel method to limit Argentine ant colonies through their natural fungal pathogens

Biofertilizers and Biocontrol Agents for Agriculture: How to Identify and Develop New Potent Microbial Strains and Traits

Microbiological tools, biofertilizers, and biocontrol agents, which are bacteria and fungi capable of providing beneficial outcomes in crop plant growth and health, have been developed for several decades. Currently we have a selection of strains available as products for agriculture, predominantly based on plant-growth-promoting rhizobacteria (PGPR), soil, epiphytic, and mycorrhizal fungi, each having specific challenges in their production and use, with the main one being inconsistency of field performance. With the growing global concern about pollution, greenhouse gas accumulation, and increased need for plant-based foods, the demand for biofertilizers and biocontrol agents is expected to grow. What are the prospects of finding solutions to the challenges on existing tools? The inconsistent field performance could be overcome by using combinations of several different types of microbial strains, consisting various members of the full plant microbiome. However, a thorough understanding of each microbiological tool, microbial communities, and their mechanisms of action must precede the product development. In this review, we offer a brief overview of the available tools and consider various techniques and approaches that can produce information on new beneficial traits in biofertilizer and biocontrol strains. We also discuss innovative ideas on how and where to identify efficient new members for the biofertilizer and biocontrol strain family

Does Intraspecific Variation in rDNA Copy Number Affect Analysis of Microbial Communities?

Amplicon sequencing of partial regions of the ribosomal RNA loci (rDNA) is widely used to profile microbial communities. However, the rDNA is dynamic and can exhibit substantial interspecific and intraspecific variation in copy number in prokaryotes and, especially, in microbial eukaryotes. As change in rDNA copy number is a common response to environmental change, rDNA copy number is not necessarily a property of a species. Variation in rDNA copy number, especially the capacity for large intraspecific changes driven by external cues, complicates analyses of rDNA amplicon sequence data. We highlight the need to (i) interpret amplicon sequence data in light of possible interspecific and intraspecific variation, and (ii) examine the potential plasticity in rDNA copy number as an important ecological factor to better understand how microbial communities are structured in heterogeneous environments.peerReviewe

Weather in two climatic regions shapes the diversity and drives the structure of fungal endophytic community of bilberry (Vaccinium myrtillus L.) fruit

Abstract Background Bilberry (Vaccinium myrtillus L.) is one of the most important economic and natural resources in Northern Europe. Despite its importance, the endophytic fungal community of the fruits has rarely been investigated. Biogeographic patterns and determinants of the fungal diversity in the bilberry fruit are poorly understood, albeit fungal endophytes can have a close relationship with the host plants. Here, we investigated the effect of climatic regions, and their weather conditions within growth season and soil properties on fungal endophytic communities of bilberry fruits collected from northern and southern regions of Finland using high-throughput sequencing technology targeting the internal transcribed spacer 2 ribosomal DNA region for fungi. Results Species richness and beta diversity (variation in community structure) were higher in the southern compared to the studied northern region. The weather condition of the growth season drove both fungal richness and community structure. Furthermore, abundance of the genera Venturia, Cladosporium, and Podosphaera was influenced by the weather, being different between the south and north regions. Conclusions We conclude that diversity and assembly structure of the fungal endophytes in bilberry fruits follow similar patterns as for foliar fungal endophytes, being shaped by various environmental factors, such as the climate and surrounding vegetation